Project Summary/Abstract Pulmonary hypertension (PH) is associated with pulmonary vascular remodeling that promotes right heart failure and premature death. In PH, pulmonary endothelial dysfunction is identified as a critical mediator of aberrant distal arteriole remodeling to induce pulmonary hypertension. The current WHO classification system segregates PH patients into distinct categories based on predisposing conditions. Despite this, it remains unknown if patients within these groups share a common endothelial or endovascular pathophenotype that is a determinant of disease progression or response to pharmacotherapy. This has led to our hypothesis that an integrated pulmonary artery endothelial (patho)phenotypic analysis will identify previously unrecognized features shared between subsets of patients with PH that will inform on disease status. To address this, we propose 3 specific aims, which are designed to 1) establish an integrated pulmonary artery endothelial profile through microRNA, mRNA, and metabolyte expression analyses using RNA-Seq and proteomics; 2) evaluate the pulmonary artery endothelial response to exercise; and 3) correlate the endothelial phenotypic profile with clinical assessments of hemodynamics, right ventricular performance and functional capacity. Studies to profile the pulmonary vascular endothelium will be performed using immediately isolated pulmonary artery endothelial cells obtained at the time of right heart catheterization and plasma samples drawn in the pre- and post- exercise phases of a 6-minute walk test. Using a systems biology approach, data from the advanced phenotypic profiling studies will be incorporated into a network to uncover previously unknown interactions between microRNAs, mRNAs, and metabolic proteins that function as disease modifiers and can be developed further as targets for pharmacotherapies or biomarkers. Integrated endothelial pathophenotypic profiling will be accomplished through collaboration of a multidisciplinary team that includes individuals with expertise in pulmonary vascular disease, cardiology, imaging, cell and molecular biology, and bioinformatics. Completion of the integrated phenotyping proposal will lead to the emergence of key endothelial phenotypes that will redefine our approach to patients with PH.